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Literature summary for 3.6.4.13 extracted from

  • Yamashita, T.; Unno, H.; Mori, Y.; Tani, H.; Moriishi, K.; Takamizawa, A.; Agoh, M.; Tsukihara, T.; Matsuura, Y.
    Crystal structure of the catalytic domain of Japanese encephalitis virus NS3 helicase/nucleoside triphosphatase at a resolution of 1.8 A (2008), Virology, 373, 426-436.
    View publication on PubMed

Application

Application Comment Organism
pharmacology conservation of the NTP-binding pocket among viruses of the family Flaviviridae as potential for development of therapeutics Japanese encephalitis virus

Cloned(Commentary)

Cloned (Comment) Organism
expressed in Escherichia coli BL21 (DE3), recombinant protein, pET21b vector Japanese encephalitis virus

Crystallization (Commentary)

Crystallization (Comment) Organism
enzymatically active fragment of the JEV NTPase/helicase catalytic domain, recombinant protein, crystal structure determined at 1.8 A resolution, data collection and refinement statistics Japanese encephalitis virus

Protein Variants

Protein Variants Comment Organism
G199A mutation in WALKER A motif, PCR-based mutagenesis, ATPase and RNA helicase activity lost Japanese encephalitis virus
G460A mutation of residues of the arginine finger within the active sites of ATP hydrolysis, no effect on either ATPase or RNA-unwinding activities Japanese encephalitis virus
G463A mutation of residues of the arginine finger within the active sites of ATP hydrolysis, no effect on either ATPase or RNA-unwinding activities Japanese encephalitis virus
K200A mutation in WALKER A motif, PCR-based mutagenesis, ATPase and RNA helicase activity lost Japanese encephalitis virus
K200D PCR-based mutagenesis, ATPase and RNA helicase activity lost Japanese encephalitis virus
K200E PCR-based mutagenesis, ATPase and RNA helicase activity lost Japanese encephalitis virus
K200H PCR-based mutagenesis, ATPase and RNA helicase activity lost Japanese encephalitis virus
K200N PCR-based mutagenesis, ATPase and RNA helicase activity lost Japanese encephalitis virus
K200Q PCR-based mutagenesis, ATPase and RNA helicase activity lost Japanese encephalitis virus
K200R PCR-based mutagenesis, ATPase and RNA helicase activity lost Japanese encephalitis virus
Q457A mutation of residues of the arginine finger within the active sites of ATP hydrolysis, 80% reduction of ATPase activity, no RNA helicase activity Japanese encephalitis virus
R458A mutation of residues of the arginine finger within the active sites of ATP hydrolysis, 90% reduction of ATPase activity, no RNA helicase activity Japanese encephalitis virus
R459A mutation of residues of the arginine finger within the active sites of ATP hydrolysis, no effect on either ATPase or RNA-unwinding activities Japanese encephalitis virus
R461A mutation of residues of the arginine finger within the active sites of ATP hydrolysis, no ATPase activity, no RNA helicase activity Japanese encephalitis virus
R464A mutation of residues of the arginine finger within the active sites of ATP hydrolysis, no ATPase activity, no RNA helicase activity Japanese encephalitis virus
T201A mutation in WALKER A motif, PCR-based mutagenesis, ATPase and RNA helicase activity lost Japanese encephalitis virus
V462A mutation of residues of the arginine finger within the active sites of ATP hydrolysis, no effect on either ATPase or RNA-unwinding activities Japanese encephalitis virus

Metals/Ions

Metals/Ions Comment Organism Structure
Mg2+ no ATPase activity of the wild-type in the absence of Japanese encephalitis virus

Molecular Weight [Da]

Molecular Weight [Da] Molecular Weight Maximum [Da] Comment Organism
54000
-
molecular mass of the helicase/NTPase domain, SDS-PAGE Japanese encephalitis virus

Natural Substrates/ Products (Substrates)

Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
ATP + H2O Japanese encephalitis virus
-
ADP + phosphate
-
?

Organism

Organism UniProt Comment Textmining
Japanese encephalitis virus P27395
-
-

Purification (Commentary)

Purification (Comment) Organism
gel filtration, recombinant protein Japanese encephalitis virus

Specific Activity [micromol/min/mg]

Specific Activity Minimum [µmol/min/mg] Specific Activity Maximum [µmol/min/mg] Comment Organism
additional information
-
structural characterization of catalytic domain, mutation analysis of residue substitution in the Walker A motif (Gly199, Lys200 and Thr201), within the NTP-binding pocket (Gln457, Arg461 and Arg464) and of Arg458 in the outside of the pocket in the motif IV, residues crucial for ATPase and RNA helicase activities and virus replication, Lys200 cannot be substituted by other residues to establish sufficient activities, structure of the NTP-binding pocket well conserved among the viruses of the Flaviviridae Japanese encephalitis virus

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
ATP + H2O
-
Japanese encephalitis virus ADP + phosphate
-
?
ATP + H2O genome structure, crystals and three-dimensional structure determined, structure of NTP-binding region, conserved residues within the NTP-binding pocket, ATPase and RNA helicase activities determined Japanese encephalitis virus ADP + phosphate
-
?

Synonyms

Synonyms Comment Organism
nonstructural protein 3 ambiguous Japanese encephalitis virus

Temperature Optimum [°C]

Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
37
-
assay at Japanese encephalitis virus